Dioxygen reduction serves two primary biological roles, energy coupling and substrate oxygenation. The heme iron O2 sensors and electron transport systems show many common properties but differ in that the latter -- mono and dioxygenases -- also bind carbon substrate and have fewer components. These oxygenases form "inert" organic groups into the main stream of metabolism for synthetic, detoxifying and carbon cycling processes. The mitochondrial and the microbial methylene systems use two specific iron proteins -- heme sulfur and sulfide -- in dienzyme complexes, regulated by substrates to perform stereo-selective substrate specific hydroxylations. The less specific, substrate inducible, microsomal detoxification monoxygenases contain only the heme sulfur protein, reduced by flavo-proteins in the presence of specific phospholipid effectors. The pure microbial methylene hydroxylase enzymes prepared in our laboratory from Pseudomonas putida have provided the proteins for structural and dynamic analyses of the sites, stable intermediates and reaction processes applicable to all P450 monoxygenase systems. Two oxygenated heme protein intermediates have now been characterized, (FeO2) plus 2 and (FeO) plus 3. The redox equilibria and binding of carbon and oxygen substrates have been analyzed in the thermodynamic parameters as has the dienzyme complex formation between the two iron proteins. Modification of selected amino acid residues and completion of the primary structure of heme monoxygenase are the 1976-7 objectives; the structure of Pd has been completed. Free mercapto groups, 6 for protein, influence stability and reactions with oxygen and carbon. The ferric heme protein reacts with peracids to form oxygenating intermediate Compound I of the peroxidases now under intensive analysis in our laboratory. Low temperature cosolvent systems are in use in collaboration with Pierre Douzou and Pascale Debey to determine the possible role of this intermediate in the full P450 hydroxylase system. BIBLIOGRAPHIC REFERENCES: Gunsalus, I.C. Thermodynamic Control Features of the Pseudomonas putida Monoxygenase System. In The Structural Basis of Membrane Function (Hatefi, Y., and Djavadi-Ohaniance, L., eds.), Academic Press: New York, 1976, pp. 377-387.